CN109536433A - Trichoderma harzianum high activity spore powder, preparation method thereof - Google Patents
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Abstract
The present invention is Trichoderma harzianum high activity spore powder, preparation method thereof, it include: to carry out trichoderma harzianum strain to expand fermentation step by step, it is produced from the preparation of level-one kind, second level kind, three-level kind, high concentration Trichoderma harzianum chlamydospore fermentation liquid is produced to using sublevel solution fermentation, then gained fermentation liquid is concentrated, and add different function additive and filler and be dried, obtain Trichoderma harzianum high activity conidia powder.The invention enables Trichoderma harzianum chlamydospore living bacteria counts to reach 2 × 109Cfu/g or more has many advantages, such as that high activity, shelf life are long, germination rate is high, rapidly adapt to soil environment.
Description
Technical field
The present invention relates to a kind of Trichoderma harzianum chlamydospore fermentation technique and high viable count spore powder, preparation method thereofs.
Background technique
Trichoderma harzianum has a variety of biological and ecological methods to prevent plant disease, pests, and erosions and growth-promoting performance, and effect is good, Environmental Safety, is increasingly welcome by market.
And Trichoderma harzianum spore has the characteristics that shelf-stable, is readily transported, is easy to apply, therefore, for realize Trichoderma harzianum commercialization,
Preparing high concentration, high activity, inexpensive Trichoderma harzianum conidia powder becomes the emphasis of formulation development.
There are two types of spore forms for Trichoderma harzianum, and one is the conidium born on mycelia side shoot, another kind is that mycelia is straight
Connect the chlamydospore of division.Currently, commercialized Trichoderma harzianum product is mostly conidium preparation, conidium in the market
It is easy culture, yield is big, but conidium anti-adversity ability is poor, and preparation shelf life is short, and Field information effect is unstable；And thick wall spore
Sub- resistance is strong, survival period is long, storage easy to process, Field information effect stability and is significantly higher than its conidium preparation, but
It is not easy to cultivate, the trichoderma harzianum chlamydospore preparation of commercialization is considerably less at home and abroad.
Common Trichoderma harzianum spore powder, preparation method thereof crushes first is that air-drying or drying Trichoderma harzianum solid culture
Afterwards, it is collected by different modes such as sieving or air classifications；Second is that directly the methods of washing, filtering, low temperature drying carry out Kazakhstan thatch
Reesei spores are collected.These collection methods respectively have advantage and disadvantage, according to first method, culture drying and dry total degree
For n, finally obtaining spore quantity compared with beginning culture reduces 10 n times sides times；According to second method, although ensure that
Product Conidia persistence, but technique is cumbersome, and drying time is longer, and energy consumption is higher, and cost is significantly increased, and weakens the market of product
Competitiveness.
Summary of the invention
It is at low cost it is an object of that present invention to provide a kind of preparation process is simple, high of the invention of product Conidia persistence
Trichoderma harzianum high activity spore powder, preparation method thereof.
Above-mentioned purpose of the present invention is achieved by the following technical programs:
A kind of Trichoderma harzianum high activity spore powder, preparation method thereof, method includes the following steps:
(1) prepared by level-one kind, is inoculated into Trichoderma harzianum on solid plate culture medium, is covered with culture dish to mycelia, is completed one
It is prepared by grade kind；
(2) prepared by second level kind, is that level-one kind is inoculated into the fluid nutrient medium prepared, shaking table culture, completes second level kind system
It is standby；
(3) prepared by three-level kind, is that second level kind is inoculated into fluid nutrient medium, fermentation tank culture, completes three-level kind and prepares；
(4) Trichoderma harzianum chlamydospore is fermented, and is that three-level kind is inoculated into chlamydospore fluid nutrient medium, fermentor is divided to two ranks
Section culture, completes Trichoderma harzianum chlamydospore fermented and cultured；
After the Trichoderma harzianum chlamydospore fermentation 24~48h of first stage culture, microelement-supplementing adjusts fermentation liquid
The liquid fermentations parameter such as pH, cultivation temperature, air inflow is completed to breathe out thatch into second stage fermented and cultured after cultivating 24~48h
Trichoderma chlamydospore fermentation.
In step (1), solid plate culture medium is Martin (Martin) culture medium, constituent are as follows: peptone 5g/
L, glucose 10g/L, potassium dihydrogen phosphate 1g/L, magnesium sulfate 0.5g/L, 1% rose-bengal aqueous solution 3.3mL/L, agar 18~
20g/L, last moisturizing are dissolved to 1L, 121 DEG C of high pressure sterilization 20min, condition of culture are as follows: cultivation temperature is 28~30 DEG C；Culture
Time is 4~7d.
In step (1), solid plate culture medium is PDA culture medium, constituent are as follows: potato 200g/L, glucose
20g/L, 18~20g/L of agar, last moisturizing are dissolved to 1L, 121 DEG C of high pressure sterilization 30min, condition of culture are as follows: cultivation temperature is
28~30 DEG C；Incubation time is 4~7d.
In step (1), solid plate culture medium is Martin's culture medium, constituent are as follows: potassium dihydrogen phosphate 1g/L, grape
Sugar 10 g/L, magnesium sulfate 0.5g/L, peptone 5 g/L, 1% rose-bengal aqueous solution 3.3mL, 18~20g/L of agar are finally mended
Water is dissolved to 1L, 121 DEG C of high pressure sterilization 20min, condition of culture are as follows: cultivation temperature is 28~30 DEG C；Incubation time is 4~7d.
Fluid nutrient medium described in step (2), (3), constituent are as follows: 10~40 g/L oatmeals, sucrose 0.5~10
G/L, 0.5~5 g/L of potassium dihydrogen phosphate, 0.5~5 g/L of magnesium sulfate, last moisturizing are dissolved to 1L, 121 DEG C of high pressure sterilizations
20min。
In step (2), it is that 5~10mm punch punches on first order seed that inoculum concentration, which is with diameter, every 100mL liquid
5~15 pieces of cultures of culture medium inoculated, condition of culture are as follows: cultivation temperature is 28~30 DEG C, 60~120rpm/min of revolving speed, culture
Time is 24~72h.
In step (3), inoculum concentration is every 1L fluid nutrient medium, by volume 5~15% inoculation second level kind；Condition of culture are as follows:
Cultivation temperature is 28~30 DEG C；60~120rpm/min of revolving speed, ventilatory capacity are 0.05~0.1MPa, and incubation time is 24~48h.
In step (4), chlamydospore fluid nutrient medium, constituent are as follows: 10~40 g/L oatmeals, 10~30 g/L
Soybean powder, 5~20 g/L of sucrose, 0.5~5 g/L of potassium dihydrogen phosphate, 3~5 kinds in 0.5~5 g/L of magnesium sulfate, rear moisturizing is fixed
It is molten to 1L, 121 DEG C of high pressure sterilization 30min, inoculum concentration is every 1L chlamydospore fluid nutrient medium, by volume 5~20% inoculation three
Grade kind.First stage condition of culture are as follows: cultivation temperature is 28~30 DEG C；60~120rpm/min of revolving speed, ventilatory capacity be 0.05~
0.1MPa, incubation time is 24~48h, and after the first stage, inorganic salts mother liquor is added in every 1L culture, so that every liter of culture
Object adds containing 1~3 kind in 0.001~0.01 g manganese chloride, 0.01~0.05 g zinc sulfate, 0.001~0.01 g iron chloride
Enter appropriate 0.5~1mol/L hydrochloric acid adjustment pH to 6~4, second stage condition of culture are as follows: cultivation temperature is 20~27 DEG C；Revolving speed
140~200rpm/min, ventilatory capacity are 0.1~0.2MPa, and incubation time is 24~48h.
Step (4) fermentation liquid is prepared into concentration liquid, using miillpore filter-filtering or miillpore filter-centrifugal dehydration
Mode be made high activity conidia powder, miillpore filter aperture select 1~10 μm.
Step (4) fermentation liquid is prepared into concentration liquid, a certain proportion of heat resistant freeze drying protection is added in concentrate
Agent, wetting and dispersing suspending agent, filler etc. carry out spray drying or cryogenic vacuum freeze-drying after mixing evenly, high activity are made
Conidia powder, the heat-resisting lyophilized protecting agent be trehalose, glucan, glycerol, polyethylene glycol, peptone, polysorbas20, glycine,
Sodium glutamate, soluble starch, gossypose, galactolipin, fructose, sucrose, 1~3 kind in dimethyl sub-maple；The wetting and dispersing
Suspending agent is sodium carboxymethylcellulose, naphthalenesulfonate formaldehyde condensation compound, sodium lignin sulfonate, polyvinyl alcohol, polysorbate85, defatted milk
1~3 kind in powder；The filler is white carbon black, in turf, kaolin, precipitated calcium carbonate, attapulgite, diatomite, talcum powder
1~2 kind.Drying process with atomizing parameter are as follows: 85~150 DEG C of inlet temperature, outlet temperature be 60~95 DEG C, frequency be 3~
12HZ；Cryogenic vacuum freeze drying process parameter are as follows: pre-freezing temperature -100~-50 DEG C, pre-freeze time are 1~5h, and thickness is lyophilized
1~3cm, 28~40h of freeze-drying time.
The present invention has the following advantages:
Preparation process is simple, at low cost, and product Conidia persistence is high, and miscellaneous bacteria rate is low, and fineness is high, can produce high-quality Trichoderma harzianum
Preparation.Trichoderma harzianum chlamydospore living bacteria count reaches 2 × 109Cfu/g or more has high activity, shelf life length, sprouts
Rate is high, rapidly adapts to the advantages that soil environment.
Specific embodiment
It is explained further the present invention with reference to embodiments, but embodiment does not do any type of limit to the present invention
It is fixed.
Embodiment 1:
1, level-one kind is prepared:
(1) it prepares PDA plate: taking 40g to peel after potato boils 30min, potato juice is obtained by filtration, glucose 4g is added,
Agar 3.6g, moisturizing is dissolved to 200mL, 121 DEG C of sterilizing 30min, after sterilizing, pours into prior sterilized culture dish, cooling
It is spare；
(2) prepared by level-one kind: under aseptic technique, Trichoderma harzianum being inoculated on PDA plate, 30 DEG C of culture 5d.
2, second level kind is prepared:
(1) preparation of culture medium: taking 70g oatmeal, 1.8g sucrose, 1.8g dipotassium hydrogen phosphate, 2.1g magnesium sulfate adds water to be settled to
3.5L, 121 DEG C of sterilizing 20min；
(2) prepared by second level kind: being 6mm punch with aperture under aseptic technique, beats on the level-one kind plate prepared
Hole is inoculated into the culture medium prepared, shaking table culture by 7 pieces/100mL inoculum concentration, and 30 DEG C of cultivation temperature, revolving speed 120rpm/
Min, 50 h of incubation time.
3, three-level kind is prepared:
(1) preparation of culture medium: taking 700g oatmeal, 18g sucrose, 17.5g dipotassium hydrogen phosphate, 21g magnesium sulfate fills after being boiled into paste
Enter 50L liquid fermentation tank, suitable quantity of water is added, makes to fill culture final volume 31.5L in fermentor after sterilization and cooling, 121 DEG C go out
Bacterium 20min；
(2) prepared by three-level kind: under aseptic technique, it is inoculated with the 3.5L second level kind prepared, 30 DEG C of cultivation temperature, and revolving speed
80rpm/min, ventilatory capacity 0.06MPa, incubation time is for 24 hours.
4, Trichoderma harzianum chlamydospore is fermented:
(1) preparation of culture medium: taking 8kg oatmeal, 5.3kg soybean powder, 3kg sucrose, 250g dipotassium hydrogen phosphate, 280g magnesium sulfate,
It is packed into 500L liquid fermentation tank after being boiled into paste, suitable quantity of water is added, makes to fill culture final volume in fermentor after sterilization and cooling
315L, 121 DEG C of sterilizing 30min；
(2) first stage ferments: under aseptic technique, it is inoculated with the 35L three-level kind prepared, 30 DEG C of cultivation temperature, and revolving speed
80rpm/min, ventilatory capacity 0.06MPa, incubation time is for 24 hours；
(3) second stage is fermented: 100g/L manganese chloride solution 4mL, 200g/L solution of zinc sulfate 18mL of addition filtration sterilization,
200g/L ferric chloride solution 15mL, and 1mol/L hydrochloric acid adjustment pH to 5.7 is added, cultivation temperature is adjusted to 21 DEG C, and revolving speed adjusts
To 160rpm/min, ventilatory capacity is adjusted to 0.17 MPa, continues to cultivate 36h；
5, fermentation liquid is concentrated:
Fermentation liquid is filtered with 10 μm of micropore filtering film, it is spare to collect concentrate.
6, prepared by high activity conidia powder:
30 g/L of glucan, 10 g/L of peptone, 50 g/L sodium carboxymethylcelluloses, the poly- second of 0.2 g/L is added in every liter of concentrate
Enol, 1 g/L polysorbate85,100 g/L of kaolin carry out cryogenic vacuum freeze-drying, Drying Technology Parameter after mixing evenly
For -98 DEG C of pre-freezing temperature, the pre-freeze time is 1.5h, and thickness 2.5cm, freeze-drying time 40h is lyophilized.Ha Ci is obtained after the completion of freeze-drying
Trichoderma high activity conidia powder, living bacteria count 4.5 × 109cfu/g。
Embodiment 2:
1, level-one kind is prepared:
(1) it prepares Martin plate: taking 1g peptone, 2g glucose, 0.2g potassium dihydrogen phosphate, 0.1g magnesium sulfate, 1% Bangladesh
Red aqueous solution 0.66mL, agar 3.6g, moisturizing are dissolved to 200mL, and 121 DEG C of sterilizing 20min after sterilizing, are poured into and sterilized in advance
Culture dish in, cooling it is spare；
(2) prepared by level-one kind: under aseptic technique, Trichoderma harzianum being inoculated on Martin plate, 28 DEG C of culture 5d.
2, second level kind is prepared:
(1) preparation of culture medium: taking 78g oatmeal, 1.4g sucrose, 1.6g dipotassium hydrogen phosphate, 6g magnesium sulfate adds water to be settled to 2L,
121 DEG C of sterilizing 30min；
(2) prepared by second level kind: being 10mm punch with aperture under aseptic technique, on the level-one kind plate prepared
Punching, by 5 pieces/100mL inoculum concentration, is inoculated into the culture medium prepared, shaking table culture, 28 DEG C of cultivation temperature, revolving speed
70rpm/min, incubation time 40h.
3, three-level kind is prepared:
(1) preparation of culture medium: taking 360g oatmeal, 8.5g sucrose, 9g dipotassium hydrogen phosphate, 7g magnesium sulfate is packed into after being boiled into paste
Suitable quantity of water is added in 20L liquid fermentation tank, makes to fill culture final volume 12L in fermentor after sterilization and cooling, 121 DEG C of sterilizings
20min；
(2) prepared by three-level kind: under aseptic technique, it is inoculated with the 2L second level kind prepared, 28 DEG C of cultivation temperature, and revolving speed
110rpm/min, ventilatory capacity 0.06MPa, incubation time is for 24 hours.
4, Trichoderma harzianum chlamydospore is fermented:
(1) preparation of culture medium: taking 1.5kg soybean powder, 2.5kg sucrose, 600g dipotassium hydrogen phosphate is packed into 200L liquid after being boiled into paste
Suitable quantity of water is added in body fermentor, makes to fill culture final volume 126L, 121 DEG C of sterilizing 30min in fermentor after sterilization and cooling；
(2) first stage ferments: under aseptic technique, it is inoculated with the 14L three-level kind prepared, 30 DEG C of cultivation temperature, and revolving speed
80rpm/min, ventilatory capacity 0.1MPa, incubation time 30h；
(3) second stage is fermented: 100 g/L manganese chloride solution 13.5mL, the 200 g/L ferric chloride solutions of filtration sterilization are added
2.7mL, and 1mol/L hydrochloric acid adjustment pH to 4.3 is added, cultivation temperature is adjusted to 27 DEG C, and revolving speed is adjusted to 150rpm/min, is led to
Tolerance is adjusted to 0.2MPa, continues to cultivate 40h；
5, fermentation liquid is concentrated:
Fermentation liquid is filtered with 8 μm of micropore filtering film, it is spare to collect concentrate.
6, prepared by high activity conidia powder:
Every liter of concentrate is added 10 g/L of sodium glutamate, 90 g/L of soluble starch, 20 g/L fructose, 5 g/L skimmed milk powers,
3 g/L sodium lignin sulfonates, naphthalenesulfonate formaldehyde condensation compound, 20 g/L of white carbon black, 70 g/L of attapulgite, after mixing evenly,
It is spray-dried, Drying Technology Parameter is 145 DEG C of inlet temperature, and outlet temperature is 95 DEG C, frequency 10HZ；Spray drying
Trichoderma harzianum high activity conidia powder, living bacteria count 2.1 × 10 are obtained after the completion9cfu/g。
Embodiment 3:
1, level-one kind is prepared:
(1) it prepares Martin plate: taking 1g peptone, 2g glucose, 0.2g potassium dihydrogen phosphate, 0.1g magnesium sulfate, 1% Bangladesh
Red aqueous solution 0.66mL, agar 3.6g, moisturizing are dissolved to 200mL, and 121 DEG C of sterilizing 20min after sterilizing, are poured into and sterilized in advance
Culture dish in, cooling it is spare；
(2) prepared by level-one kind: under aseptic technique, Trichoderma harzianum being inoculated on Martin plate, 28 DEG C of culture 5d.
2, second level kind is prepared:
(1) preparation of culture medium: taking 9g oatmeal, 7g sucrose, 0.35g dipotassium hydrogen phosphate, 0.04g magnesium sulfate adds water to be settled to
0.7L, 121 DEG C of sterilizing 30min；
(2) prepared by second level kind: being 6mm punch with aperture under aseptic technique, beats on the level-one kind plate prepared
Hole is inoculated into the culture medium prepared, shaking table culture by 8 pieces/100mL inoculum concentration, and 29 DEG C of cultivation temperature, revolving speed 90rpm/
Min, incubation time 72h.
3, three-level kind is prepared:
(1) preparation of culture medium: taking 167g oatmeal, 130g sucrose, 10g dipotassium hydrogen phosphate, 52g magnesium sulfate fills after being boiled into paste
Enter 20L liquid fermentation tank, suitable quantity of water is added, makes to fill culture final volume 13L in fermentor after sterilization and cooling, 121 DEG C of sterilizings
20min；
(2) prepared by three-level kind: under aseptic technique, it is inoculated with the 0.7L second level kind prepared, 29 DEG C of cultivation temperature, and revolving speed
90rpm/min, ventilatory capacity 0.08MPa, incubation time 40h.
4, Trichoderma harzianum chlamydospore is fermented:
(1) preparation of culture medium: taking 4.8kg oatmeal, 560g dipotassium hydrogen phosphate, 76g magnesium sulfate is packed into 200L liquid after being boiled into paste
Suitable quantity of water is added in body fermentor, makes to fill culture final volume 127L, 121 DEG C of sterilizing 30min in fermentor after sterilization and cooling；
(2) first stage ferments: under aseptic technique, it is inoculated with the 13L three-level kind prepared, 29 DEG C of cultivation temperature, and revolving speed
90rpm/min, ventilatory capacity 0.8MPa, incubation time 40h；
(3) second stage is fermented: 200 g/L solution of zinc sulfate 30mL, the 200 g/L ferric chloride solutions of filtration sterilization are added
2mL, and 0.8mol/L hydrochloric acid adjustment pH to 5.9 is added, cultivation temperature is adjusted to 22 DEG C, and revolving speed is adjusted to 200rpm/min, is led to
Tolerance is adjusted to 0.17MPa, continues to cultivate 28h；
5, fermentation liquid is concentrated:
Fermentation liquid is handled with 6 μm of micropore filtering film centrifugal dehydration, it is spare to collect concentrate.
6, prepared by high activity conidia powder:
Every liter of concentrate is added 50 g/L of glycine, 90 g/L of gossypose, 0.3 g/L polyvinyl alcohol, 40 g/L skimmed milk powers,
80 g/L of diatomite, 10 g/L of precipitated calcium carbonate are spray-dried after mixing evenly, and Drying Technology Parameter is import temperature
90 DEG C of degree, outlet temperature are 65 DEG C, frequency 4HZ；Trichoderma harzianum high activity conidia powder is obtained after the completion of spray drying, it is effectively living
Bacterium number 3.2 × 109cfu/g。
Claims (10)
1. a kind of Trichoderma harzianum high activity spore powder, preparation method thereof, method includes the following steps:
(1) prepared by level-one kind, is inoculated into Trichoderma harzianum on solid plate culture medium, is covered with culture dish to mycelia, is completed one
It is prepared by grade kind；
(2) prepared by second level kind, is that level-one kind is inoculated into the fluid nutrient medium prepared, shaking table culture, completes second level kind system
It is standby；
(3) prepared by three-level kind, is that second level kind is inoculated into fluid nutrient medium, fermentation tank culture, completes three-level kind and prepares；
(4) Trichoderma harzianum chlamydospore is fermented, and is that three-level kind is inoculated into chlamydospore fluid nutrient medium, fermentor point the
One, the two-stage cultivates, and completes Trichoderma harzianum chlamydospore fermented and cultured；
After the Trichoderma harzianum chlamydospore fermentation first stage cultivates 24~48h, microelement-supplementing, adjustment fermentation liquid pH,
The liquid fermentations parameter such as cultivation temperature, air inflow is completed to breathe out thatch wood into second stage fermented and cultured after cultivating 24~48h
Mould chlamydospore fermentation.
2. the Preparation Method of system according to claim 1, which is characterized in that in step (1), solid plate culture medium is Martin
Family name (Martin) culture medium, constituent are as follows: peptone 5g/L, glucose 10g/L, potassium dihydrogen phosphate 1g/L, magnesium sulfate
0.5g/L, 1% rose-bengal aqueous solution 3.3mL/L, 18~20g/L of agar, last moisturizing are dissolved to 1L, 121 DEG C of high pressure sterilizations
20min, condition of culture are as follows: cultivation temperature is 28~30 DEG C；Incubation time is 4~7d.
3. the Preparation Method of system according to claim 1, which is characterized in that in step (1), solid plate culture medium is PDA
Culture medium, constituent are as follows: potato 200g/L, glucose 20g/L, 18~20g/L of agar, last moisturizing are dissolved to 1L,
121 DEG C of high pressure sterilization 30min, condition of culture are as follows: cultivation temperature is 28~30 DEG C；Incubation time is 4~7d.
4. the Preparation Method of system according to claim 1, which is characterized in that in step (1), solid plate culture medium is Martin
Culture medium, constituent are as follows: potassium dihydrogen phosphate 1g/L, glucose 10 g/L, magnesium sulfate 0.5g/L, 5 g/L of peptone, 1% Meng
Add and draw red aqueous solution 3.3mL, 18~20g/L of agar, last moisturizing is dissolved to 1L, 121 DEG C of high pressure sterilization 20min, condition of culture
Are as follows: cultivation temperature is 28~30 DEG C；Incubation time is 4~7d.
5. the Preparation Method of system according to claim 1, which is characterized in that fluid nutrient medium described in step (2), (3),
Constituent are as follows: 10~40 g/L oatmeals, 0.5~10 g/L of sucrose, 0.5~5 g/L of potassium dihydrogen phosphate, magnesium sulfate 0. 5~
5 g/L, last moisturizing are dissolved to 1L, 121 DEG C of high pressure sterilization 20min.
6. the Preparation Method of system according to claim 5, which is characterized in that in step (2), inoculum concentration be with diameter be 5~
10mm punch punches on first order seed, and every 100mL fluid nutrient medium is inoculated with 5~15 pieces of cultures, condition of culture are as follows: training
Supporting temperature is 28~30 DEG C, 60~120rpm/min of revolving speed, and incubation time is 24~72h.
7. the Preparation Method of system according to claim 5, which is characterized in that in step (3), inoculum concentration is every 1L Liquid Culture
Base is inoculated with 5~15% second level kind by volume；Condition of culture are as follows: cultivation temperature is 28~30 DEG C；60~120rpm/ of revolving speed
Min, ventilatory capacity are 0.05~0.1MPa, and incubation time is 24~48h.
8. the Preparation Method of system according to claim 1, which is characterized in that in step (4), the chlamydospore Liquid Culture
Base, constituent are as follows: 10~40 g/L oatmeals, 10~30 g/L soybean powders, 5~20 g/L of sucrose, potassium dihydrogen phosphate 0.5
3~5 kinds in~5 g/L, 0.5~5 g/L of magnesium sulfate, rear moisturizing is dissolved to 1L, 121 DEG C of high pressure sterilization 30min, and inoculum concentration is
Every 1L chlamydospore fluid nutrient medium is inoculated with 5~20% three-level kind, first stage condition of culture are as follows: cultivation temperature by volume
It is 28~30 DEG C；60~120rpm/min of revolving speed, ventilatory capacity are 0.05~0.1MPa, and incubation time is 24~48h, first stage
After, inorganic salts mother liquor is added in every 1L culture so that every liter of culture containing 0.001~0.01 g manganese chloride, 0.01~
1~3 kind in 0.05 g zinc sulfate, 0.001~0.01 g iron chloride, and appropriate 0.5~1mol/L hydrochloric acid adjustment pH to 6 is added
~4, second stage condition of culture are as follows: cultivation temperature is 20~27 DEG C；140~200rpm/min of revolving speed, ventilatory capacity be 0.1~
0.2MPa, incubation time are 24~48h.
9. the Preparation Method of system according to claim 1, which is characterized in that step (4) fermentation liquid is prepared into concentration
High activity conidia powder, the choosing of miillpore filter aperture is made in liquid by the way of miillpore filter-filtering or miillpore filter-centrifugal dehydration
With 1~10 μm.
10. the Preparation Method of system according to claim 1, which is characterized in that step (4) fermentation liquid is prepared into concentration
A certain proportion of heat-resisting lyophilized protecting agent, wetting and dispersing suspending agent, filler etc. are added in concentrate, stirs evenly laggard for liquid
High activity conidia powder is made in row spray drying or cryogenic vacuum freeze-drying, and the heat-resisting lyophilized protecting agent is trehalose, Portugal is poly-
Sugar, glycerol, polyethylene glycol, peptone, polysorbas20, glycine, sodium glutamate, soluble starch, gossypose, galactolipin, fructose,
1~3 kind in sucrose, dimethyl sub-maple；The wetting and dispersing suspending agent is sodium carboxymethylcellulose, naphthalene sulphonate formaldehyde condensation
Object, sodium lignin sulfonate, polyvinyl alcohol, polysorbate85,1~3 kind in skimmed milk power；The filler is white carbon black, turf, kaolinite
Soil, precipitated calcium carbonate, attapulgite, diatomite, 1~2 kind in talcum powder, drying process with atomizing parameter are as follows: inlet temperature 85
~150 DEG C, outlet temperature is 60~95 DEG C, and frequency is 3~12HZ；Cryogenic vacuum freeze drying process parameter are as follows: pre-freezing temperature-
100~-50 DEG C, the pre-freeze time is 1~5h, and 1~3cm of thickness, 28~40h of freeze-drying time is lyophilized.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110810174A (en) * | 2019-11-15 | 2020-02-21 | 山东省科学院生态研究所(山东省科学院中日友好生物技术研究中心) | Application of trichoderma harzianum LTR-2 in cereal crop cultivation |
| CN113215074A (en) * | 2021-04-16 | 2021-08-06 | 上海大井生物工程有限公司 | Production method of trichoderma chlamydospore |
| CN115851453A (en) * | 2022-12-07 | 2023-03-28 | 中国林业科学研究院森林生态环境与自然保护研究所 | Culture medium for preparing Isaria lineare fungal spore powder and preparation method of Isaria lineare fungal spore powder |
| WO2024068640A1 (en) * | 2022-09-26 | 2024-04-04 | Biovirid B.V. | Method for producing fungal spores, liquid medium used therein and substance produced thereby |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103589646A (en) * | 2012-08-13 | 2014-02-19 | 北京中农广袤农业科技有限公司 | Trichoderma strain and purpose thereof, and chlamydospore and preparation method thereof |
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2018
- 2018-12-25 CN CN201811587186.XA patent/CN109536433A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103589646A (en) * | 2012-08-13 | 2014-02-19 | 北京中农广袤农业科技有限公司 | Trichoderma strain and purpose thereof, and chlamydospore and preparation method thereof |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110810174A (en) * | 2019-11-15 | 2020-02-21 | 山东省科学院生态研究所(山东省科学院中日友好生物技术研究中心) | Application of trichoderma harzianum LTR-2 in cereal crop cultivation |
| CN110810174B (en) * | 2019-11-15 | 2021-12-24 | 山东省科学院生态研究所(山东省科学院中日友好生物技术研究中心) | Application of trichoderma harzianum LTR-2 in cereal crop cultivation |
| CN113215074A (en) * | 2021-04-16 | 2021-08-06 | 上海大井生物工程有限公司 | Production method of trichoderma chlamydospore |
| WO2024068640A1 (en) * | 2022-09-26 | 2024-04-04 | Biovirid B.V. | Method for producing fungal spores, liquid medium used therein and substance produced thereby |
| CN115851453A (en) * | 2022-12-07 | 2023-03-28 | 中国林业科学研究院森林生态环境与自然保护研究所 | Culture medium for preparing Isaria lineare fungal spore powder and preparation method of Isaria lineare fungal spore powder |
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